Ischemic cardiomyopathy and myocardial infarction (MI) are typified by the irreversible loss of cardiac muscle (cardiomyocytes) and vasculature (composed of endothelial cells and smooth muscle cells), which are essential for maintaining cardiac integrity and function. The recent identification of adult and embryonic stem cells has triggered attempts to directly repopulate these tissues by stem cell transplantation as a novel therapeutic option. Reports describing provocative and hopeful examples of myocardial regeneration with adult stem/progenitor cells have furthered enthusiasm for the use of these cells, however many unanswered questions remain regarding their therapeutic potential and, more specifically the mechanisms responsible for the observed therapeutic effects. First, although various adult BM-derived stem cell populations have been employed in an attempt to regenerate myocardium, currently no human adult stem cells have been convincingly shown to have the capacity to induce both a therapeutic level of neovascularization and cardiomyogenesis, two of the key components for successful myocardial tissue regeneration. Second, there has been a degree of discrepancy between the therapeutic effect demonstrated in vivo and the magnitude of adult stem/progenitor (trans)differentiation into specific cell lineages. Third, although chronic post-MI heart failure is one of the most clinically relevant candidates for stem cell therapy, this therapeutic approach has been virtually uninvestigated. We recently identified a novel SC population within adult human and rat bone marrow (BM), which can be isolated at a single cell level and can be expanded for more than 120 population doublings (PDs)(>9 consecutive months). These stem cells demonstrated a potential for differentiation into cells of all three germ layers;mesoderm(endothelium, smooth muscle cells and cardiomyocytes), endoderm and neuroectoderm. We refer to these cells as BM-derived multipotent stem cells (BMSCs). Accordingly we hypothesize that transplantation of human BMSC (hBMSC) may improve cardiac function in animal models of ischemic heart diseases and the mechanisms may encompass not only cellular differentiation but also delivery of multiple humoral factors. In Specific Aim 1, we will explore the therapeutic impact of myocardial hBMSC transplantation in acute and chronic models of ischemic heart failure, which are the two most clinically relevant candidates for SC therapy. In Specific Aim 2, using cardiac samples obtained from the above experiments, we will explore local cellular and paracrine mechanisms which can mediate the therapeutic effect of hBMSC transplantation. More specifically, this will encompass cellular differentiation from hBMSC, fusion between hBMSC and host myocardium, expression of muitiple paracrine factors, and proliferative and pro-survival effects on host myocardium. In SpecificAim 3, we will investigate the systemic effect of hBMSC transplantation. Specifically,we will examine whether myocardial hBMSC transplantation can induce mobilization and recruitment of host circulating stem/progenitor cells and furthermore, explore the possibility of augmenting this natural process with the use of cytokines, thereby enhancing therapeutic efficacy. We anticipate that the results of the experiments outlined in this Proposal will yield new insight into the application of novel BM-derived human stem cells for regeneration of damaged myocardium, and potentially suggest novel therapeutic strategies for acute and chronic forms of heart failure.